blocking for western blot

Baker Matthew mbaker at cmtech.co.uk
Thu Apr 20 06:48:10 EST 2000


When using antibodies in blots that appear to give high backgound signals I
prefer to use 2-3% BSA as a blocking reagent.  It is also worth noting that
low fat (or fat free) milk powder seems to work best if used in a Western
block buffer.


Matthew
 

-----Original Message-----
From: behrends at plexus.uke.uni-hamburg.de
[mailto:behrends at plexus.uke.uni-hamburg.de]
Sent: Thursday, April 20, 2000 10:34 AM
To: methods at hgmp.mrc.ac.uk
Subject: blocking for western blot



Dear netters, 

we are routinely using milk powder as
blocking agent for western blots using 
nitrocellulose blots. However, 
we have noticed that when we leave out
milk powder altogether there is no difference. 

We now want to try products like super signal
from pierce where the company argues that you
get stronger and better signals as compared to
regular ECL solution. After our first experiments
we see that background really becomes a problem 
with this solution and that we need a good 
blocking agent. 

Since we could leave out the milk powder in our 
protocol without any problems, we start to question
whether it has any effect at all. Pierce offers 
different kinds of blocking agents, but they are
expensive and available only in large quantities. 

We would be glad to hear about any tricks or hints 
and whether the commercial solutions compare favorably
with 'home-made'-recipees. 

Thanks for your time and help
Soenke


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